Abstract

Background

Influenza A virus (IAV) neuraminidase (NA) cleaves sialic acids (Sias) from glycans.
Inhibiting NA with oseltamivir suppresses both viral infection, and viral release
from cultured human airway epithelial cells. The role of NA in viral exit is well
established: it releases budding virions by cleaving Sias from glycoconjugates on
infected cells and progeny virions. The role of NA in viral entry remains unclear.
Host respiratory epithelia secrete a mucus layer rich in heavily sialylated glycoproteins;
these could inhibit viral entry by mimicking sialylated receptors on the cell surface.
It has been suggested that NA allows influenza to penetrate the mucus by cleaving
these sialylated decoys, but the exact mechanism is not yet established.

Methods

We tested IAV interaction with secreted mucus using frozen human trachea/bronchus
tissue sections, and bead-bound purified human salivary mucins (HSM) and purified
porcine submaxillary mucins (PSM). The protective effect of mucus was analyzed using
MDCK cells coated with purified HSM and PSM with known Sia content. Oseltamivir was
used to inhibit NA activity, and the fluorescent reporter substrate, 4MU-Neu5Ac, was
used to quantify NA activity.

Results

IAV binds to the secreted mucus layer of frozen human trachea/bronchus tissues in
a Sia dependent manner. HSM inhibition of IAV infection is Sia dose-dependent, but
PSM cannot inhibit infection of underlying cells. HSM competitively inhibits NA cleavage
of 4MU-Neu5Ac, reporter substrate. Human IAV effectively cleaves Sias from HSM but
not from PSM, and binds to HSM but not to PSM.

Conclusion

IAV interacts with human mucus on frozen tissue sections and mucus-coated beads. Inhibition
of IAV infection by sialylated human mucus is dose-dependent, and enhanced when NA
is inhibited with oseltamivir. Thus NA cleaves sialylated decoys during initial stages
of infection. Understanding IAV interactions with host mucins is a promising new avenue
for drug development.